Hui T Y, Frohnert B I, Smith A J, Schaffer J E, Bernlohr D A
Department of Biochemistry, University of Minnesota, St. Paul, Minnesota 55108, USA.
J Biol Chem. 1998 Oct 16;273(42):27420-9. doi: 10.1074/jbc.273.42.27420.
Fatty acid transport protein (FATP) was identified by expression cloning strategies (Schaffer, J. E., and Lodish, H. F. (1994) Cell 79, 427-436) and shown by transfection analysis to catalyze the transfer of long-chain fatty acids across the plasma membrane of cells. It is expressed highly in tissues exhibiting rapid fatty acid metabolism such as skeletal muscle, heart, and adipose. FATP mRNA levels are down-regulated by insulin in cultured 3T3-L1 adipocytes and up-regulated by nutrient depletion in murine adipose tissue (Man, M. Z., Hui, T. Y., Schaffer, J. E., Lodish, H. F., and Bernlohr, D. A. (1996) Mol. Endocrinol. 10, 1021-1028). To determine the molecular mechanism of insulin regulation of FATP transcription, we have isolated the murine FATP gene and its 5'-flanking sequences. The FATP gene spans approximately 16 kilobases and contains 13 exons, of which exon 2 is alternatively spliced. S1 nuclease and RNase protection assays revealed the presence of multiple transcription start sites; the DNA sequence upstream of the predominant transcription start sites lacks a typical TATA box. By transient transfection assays in 3T3-L1 adipocytes, the inhibitory action of insulin on FATP transcription was localized to a cis-acting element with the sequence 5'-TGTTTTC-3' from -1347 to -1353. This sequence is very similar to the insulin response sequence found in the regulatory region of other genes negatively regulated by insulin such as those encoding phosphoenolpyruvate carboxykinase, tyrosine aminotransferase, and insulin-like growth factor-binding protein 1. Fluorescence in situ hybridization analysis revealed that the murine FATP gene is localized to chromosome 8, band 8B3.3. Interestingly, this region of chromosome 8 contains a cluster of three other genes important for fatty acid homeostasis, lipoprotein lipase, the mitochondrial uncoupling protein 1 (UCP1) and sterol regulatory element-binding protein 1. These results characterize the murine FATP gene and its insulin responsiveness as well as present a framework for future studies of its role in lipid metabolism, obesity, and type II diabetes mellitus.
脂肪酸转运蛋白（FATP）是通过表达克隆策略鉴定出来的（Schaffer, J. E., 和 Lodish, H. F. (1994) 《细胞》79, 427 - 436），转染分析表明它能催化长链脂肪酸跨细胞的质膜转运。它在脂肪酸代谢迅速的组织如骨骼肌、心脏和脂肪组织中高度表达。在培养的3T3 - L1脂肪细胞中，胰岛素可下调FATP mRNA水平，而在小鼠脂肪组织中，营养物质缺乏则会上调其水平（Man, M. Z., Hui, T. Y., Schaffer, J. E., Lodish, H. F., 和 Bernlohr, D. A. (1996) 《分子内分泌学》10, 1021 - 1028）。为了确定胰岛素调节FATP转录的分子机制，我们分离了小鼠FATP基因及其5'侧翼序列。FATP基因跨度约16千碱基，包含13个外显子，其中外显子2存在可变剪接。S1核酸酶和RNA酶保护分析揭示了多个转录起始位点的存在；主要转录起始位点上游的DNA序列缺乏典型的TATA框。通过在3T3 - L1脂肪细胞中的瞬时转染分析，胰岛素对FATP转录的抑制作用定位于一个顺式作用元件，其序列为5'-TGTTTTC-3'，位于 - 1347至 - 1353处。该序列与在其他受胰岛素负调控的基因（如编码磷酸烯醇丙酮酸羧激酶、酪氨酸转氨酶和胰岛素样生长因子结合蛋白1的基因）的调控区域中发现的胰岛素反应序列非常相似。荧光原位杂交分析表明，小鼠FATP基因定位于8号染色体8B3.3带。有趣的是，8号染色体的这个区域还包含另外三个对脂肪酸稳态很重要的基因簇，即脂蛋白脂肪酶、线粒体解偶联蛋白1（UCP1）和固醇调节元件结合蛋白1。这些结果描述了小鼠FATP基因及其胰岛素反应性，并为其在脂质代谢、肥胖和II型糖尿病中的作用的未来研究提供了一个框架。
